Chronic infection with the hepatitis C virus (HCV) is a major cause of cirrhosis, decompensated liver disease and hepatocellular carcinoma (HCC) worldwide. Globally there are an estimated 180 million persons with HCV. In the United States (U.S.), 1% of the general population or 2.7 million persons have chronic infection based on the detection of HCV RNA in serum. In 2014, chronic HCV infection was the leading cause of death in the U.S. due to an infectious cause. These figures underscore the magnitude and impact that chronic HCV infection has on global and US public health. The natural history of chronic HCV infection has been difficult to study. The protracted and silent course of infection, the absence of large cohorts of persons known to be infected, and the wide variability in outcome are major obstacles to natural history studies. Five to twenty-five percent of HCV-infected persons will develop cirrhosis over a 25-30 year period but some patients remain asymptomatic, without significant liver disease for many decades if not for life. Knowledge of the rate of progression among individuals who have not developed cirrhosis is unknown. An equally important and related issue is the clinical assessment of disease severity. Unfortunately, there are no good laboratory markers of disease severity and liver biopsy, the accepted gold standard for assessing disease severity is imperfect. Non-invasive methods to assess disease severity are highly desirable for the clinicians diagnostic toolbox. The optimal treatment for chronic HCV infection is undergoing a paradigm change with the advent of direct acting antiviral (DAA) agents. For most genotypes response rates to treatment now approach 90-95%. The challenging group of patients to treat are those with genotype 3 infection who have failed a prior course of peginterferon and ribavirin and who have advanced disease. Therapeutic options for these subjects are improving but few studies have been conducted in these patient populations. Why these subjects fail therapy is unknown. Hypotheses/problems addressed: 1) Define the host, viral and environmental factors that determine the natural history and outcome of HCV infection. To address this problem, we have created a large database of untreated patients with chronic HCV (n700) and have analyzed this database to identify factors that affect the natural history of chronic HCV infection. We have published 4 studies on this cohort, identifying clinical factors and candidate genes associated with fibrosis progression. Autoimmune hepatitis sometimes occurs in patients with chronic hepatitis C but presents difficult diagnostic and management challenges. To address the former issue we used the database to identify patients with co-existent autoimmune hepatitis and chronic hepatitis C to identify clinical and laboratory characteristics that can distinguish them from patients with chronic hepatitis C alone. Among patients with chronic hepatitis C, the findings of a positive ANA, high serum ALT, globulin and HCV levels were associated with underlying AIH. This work has been submitted for publication. We plan to assemble a cohort of treated patients with all stages of liver disease and monitor them prospectively over a 5-year period to identify the incidence and predictors of clinical outcomes. A focus of this study would be to identify predictors of outcomes after virological eradication in particular hepatocellular carcinoma and regression of liver fibrosis. The factors associated with hepatic decompensation among patients with advanced chronic hepatitis C are not well understood. Recently, it was reported that the prevalence of hepatitis E was 21% among the U.S. population. In endemic regions, hepatitis E is associated with acute on chronic liver failure. Thus, we investigated the role of hepatitis E in hepatic decompensation in the U.S. We retrospectively tested samples from patients with and without hepatic decompensation enrolled in a former study-the HALT-C study. Both the prevalence and incidence of hepatitis E was similar in patients with and without hepatic decompensation suggesting that hepatitis E does not contribute to decompensation among patients with hepatitis C in the U.S. This work was published in Clin Gastroenterol Hepatol. 2016;14(6):896-902. Finally, we are validating the usefulness of a new technology, ultrasound elastography, to non-invasively assess hepatic fibrosis. Results revealed that the device has a 96% accuracy for prediction of cirrhosis. Studies are ongoing to compare transient elastography with other non-invasive markers of fibrosis. Our goal is to develop a series of blood and imaging test that will obviate the need for liver biopsy in most patients with chronic HCV infection. 2) Develop and evaluate novel, safer and more effective therapies for chronic viral hepatitis. Management options for non-responders to peginterferon and ribavirin are limited. At the LDB, we have explored multiple options including, low dose, long term peginterferon, peginterferon and ribavirin plus SAMe, and peginterferon in combination with high dose ribavirin all with limited success. We have recently initiated a trial evaluating a once per day, pangenotypic regimen consisting of sofosbuvir and velpatasvir for a duration of twelve weeks. Seventy subjects have been enrolled and the study is ongoing. This combination of agents if efficacious, would allow patients with all genotypes to be treated with the same regimen. 3. Elucidate the viral pathogenesis of chronic HCV infection and mechanisms of action of anti-viral therapy The conduct of clinical trials over the last 30 years has allowed the LDB to acquire invaluable clinical material (patient serum, liver tissue and lymphophocytes) to which state of the art laboratory techniques can be applied to address issues of the pathogenesis of HCV infection and the mechanisms of action of antiviral therapy. In association with a clinical study using asunaprevir and daclatasvir we are seeking to answer three critical questions surrounding therapy of chronic hepatitis C: (1) what happens to the expression of interferon stimulated genes (ISG) following suppression of viral replication (2) can interferon responsiveness be restored in non-responders after suppression of viral replication and (3) why do patients with cirrhosis respond less compared to those without cirrhosis. To address these questions, we are comparing the relative expression level of ISGs before and after suppression of viremia with potent direct acting antiviral agents and focusing on genes that regulate the innate immune response. For the first question, we have shown that the level of ISGs decline rapidly, within 2 weeks following suppression of viral replication. Moreover the magnitude of decrease in ISGs was greater in patients who ultimately responded to therapy compared to patients who experienced virological breakthrough. This work was presented at the American Association for the Study of Liver Diseases meeting in San Francisco 2015 and a manuscript is ready for submission. We also explored the role of microRNAs in viral clearance during therapy with DAAs. We found 14 miRNAs that were significantly negatively correlated with ISGs between paired biopsies: miR-222, and 13 others. 13/14 miRNAs had known ISGs as targets and 2/14 target Toll like receptor. Interestingly, 9/14 miRNAs were significantly lower at baseline in responders compared to non-responders and increased at week 2 or 4 of treatment suggesting regulation by HCV. These novel results suggest that miRNAs may directly and indirectly regulate innate immunity and ISGs production and contribute to viral clearance during DAA therapy. This work was accepted for presentation at the American Association for the Study of Liver Diseases meeting in Boston in Novem